Light-activated self-sanitizing surface coatings to prevent cross-contamination from zone I and zone II surfaces

LIGHT-ACTIVATED SELF-SANITIZING SURFACE COATINGS TO PREVENT CROSS-CONTAMINATION FROM ZONE I AND ZONE II SURFACES

Sponsoring Institution

National Institute of Food and Agriculture

Project Status

ACTIVE

Funding Source

AFRI COMPETITIVE GRANT

Reporting Frequency

Annual

Accession No.

1027808

Grant No.

2022-67017-36308

Cumulative Award Amt.

$604,900.00

Proposal No.

2021-08191

Multistate No.

(N/A)

Project Start Date

Apr 1, 2022

Project End Date

Mar 31, 2026

Grant Year

2022

Program Code

[A1332]- Food Safety and Defense

Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671

Performing Department
Food Science & Technology

Non Technical Summary
Cross-contamination of food with pathogens, including bacteria and viruses, is a leading cause of food borne illnesses in the nation. Despite the implementation of good agricultural practices, good manufacturing practices, and hygiene plans, significant cross-contamination risks from the food contact surfaces (zone I) and adjoining surfaces (zone II) exist during food harvesting, processing, and food service industries. To address these challenges, the proposed research plan aims to reduce the risk of cross-contamination using food grade photoactive materials for continuous sanitation of zone I and zone II surfaces during handling and processing of food. The specific objectives are to (a) develop and evaluate food-grade photoactive antimicrobial coatings for model zone I and zone II surfaces for reducing the risk of cross-contamination; (b) evaluate the effectiveness of affinity enabled photoactive antimicrobial coatings and photoactive antimicrobial-antimicrobial "slippery" coatings to reduce the risk of cross-contamination, and (c) evaluate mechanisms of inactivation of target microbes using photo-activated coatings and assess translation of the coatings to pilot-scale systems. These food-grade photoactive coating compositions will be designed to inactivate pathogens in the presence of daylight or sunlight, thus targeting both indoor and outdoor environments. The key innovations for reducing cross-contamination are the discovery and application of food grade biopolymers coatings with endogenous photoactivity, engineering photoactive coatings using a combination of food grade compounds, bio-affinity ligands and biopolymers and development of antifouling and antimicrobial coatings. Overall, this research addresses the key unmet challenges in the areas of food safety by reducing the risk of cross-contamination.

Animal Health Component

25%

Research Effort Categories

Basic

75%

Applied

25%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
712	4099	2020	75%
712	5010	2000	25%

Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
4099 - Microorganisms, general/other; 5010 - Food;

Field Of Science
2020 - Engineering; 2000 - Chemistry;

Keywords

photoactive materials

Goals / Objectives
Overall, the proposed research plan aims to address a significant unmet need to reduce the risk of cross-contamination using food grade photoactive materials developed for applications on zone I and zone II surfaces.The overall objectives are:Aim 1: Develop and evaluate food grade photoactive antimicrobial coatings for model zone I and zone II surfaces for reducing the risk of cross-contamination from both bacterial and viral pathogens and bacterial biofilmsAim 2: Evaluate enhancement in the efficacy of the modified coatings to reduce the risk of cross-contamination by developing affinity enabled photoactive antimicrobial coatings and photoactive antimicrobial "slippery" coatings with an additional antifouling functionalityAim 3: Evaluate mechanisms of inactivation of target microbes using photo-activated coatings and assess translation of the coatings to pilot scale systems.Success in this proposal will address a long-standing unmet need in the food industry to achieve continuous sanitation of diversity of zone I and zone II surfaces using food grade photoactive coatings and reduce the risk of cross- contamination.

Project Methods
Aim 1: The experimental approach will focus on the following sub-tasks: (a) Coating of photoactive compositions of zone Iand zone II surfaces.Based on our preliminary results biopolymerswill be selected as an endogenous photo-active coating material. Chitosan modified with food-grade photoactive compounds will be also selected as a carbohydrate-based photoactive coating material. (b) The coatings will be characterized by a range of physicochemical methods including gravimetric (thermogravimetric analysis), morphological (scanning electron microscopy, SEM), physical (contact angle) and chemical analysis (Fourier-transform infrared spectroscopy, FTIR). Coating compositions will also be analyzed for mechanical durability and(c) Both the photoactivity of the coating and the antimicrobial properties of the photoactive coatings will be characterized using a combination of biochemical measurements to quantify generation of reactive oxygen species with microbiological measurements for the inactivation of inoculated bacteria and viruses.Aim 2: The experimental methods will include: coating modification with bioaffinity ligands to enhance the rate of inactivation of bacteria. To characterize the impact of this modification, the research plan will investigate the binding of the bacteria and viruses to the modified coated surface. These modified coatings will also be characterized using a combination of physico-chemical and antimicrobial activity tests as described in aim 1. Complementary to this approach, the coatings will be modified with slippery functionality. In this approach,novel materials using a combination of nanofibers and photoactive materials. Since these compositions are not yet approved for food contact applications, this research plan will focus on zone II surfaces. In this research, cellulose nanofibers (CNFs) will be functionalized with the fluoro silane to create a hydrophobic surface coating. Well-dispersed native hydrophilic CNF (1 wt %) will be mixed with a fluoro-silane under vigorous stirring conditions and kept for 6−7 h at room temperature. The physico-chemical and antimicrobial properties of the slippery coatings will also be evaluated.Aim 3:The methods in aim 3 will focus on (a)Characterization of biological damages induced by photoactive coatings: The analysis will focus on following biological damages in bacteria-induced by photoactive coatings.Analysis of membrane damage: Membrane damage will be analyzed by using the fluorescent probe propidium iodide (PI) and scanning electron microscopy (SEM).Cellular redox potential: We will use the Thiol Detection Assay Kit to measure the free thiol content in bacteria after treatments.Intracellular ATP levels: Changes in the intracellular ATP levels will be measured using the standard ATP assay kit before and after processing.DNA damage: DNA damage will be assessed based on the generation of free 3′-OH DNA ends, produced by either direct damage or excision DNA repair using the TUNEL assay.Viral damage analysis: Transmission Electron Microscopy (TEM) will be used to determine any structural or capsid damage of the treated viruses (HAV and TV) compared to controls as described in our earlier studies. SDS-PAGE will be used to determine changes to the capsid protein of the treated viruses compared to their untreated controls using previously described protocols. RNA degradation can be determined by long-template RT-PCR as only undamaged intact long RNA will be amplified.Gene expression analysis: To characterize changes in gene expression we will select a strain of E. coli K-12 MG1655, as genomic sequence data for this strain is publicly available. After treatment with optimized coating compositions in Aims 1 and 2, bacterial cultures will be stabilized with RNA protect Bacteria Reagent.The expression of 30 genes (membrane synthesize genes, membrane stress response genes, cellular redox metabolism, and oxidative stress response genes) will be examined by RT-qPCR. These genes will be selected based on the database of E. coli responses to oxidative, metabolic, and membrane damage stresses (https://ecocyc.org). In addition to this analysis, two pilot-scale studies focusing on cross-contamination reduction in pilot plant and food preparation environments will be evaluated.

Progress 04/01/23 to 03/31/24

Outputs
Target Audience:The target audience includes researchers at universities, government, and food industries. In addition, the technologies developed through this project are of broader interest to food companies for the translation of research from the lab to industrial practice. Changes/Problems:Due to the successful placement of a postdoctoral scholar in a faculty position, I had to recruit another postdoctoral fellow for this project. Due to a lag in joining the postdoctoral scholar, we were behind by a few months. In addition, I am working with the former postdoctoral fellow to complete the pending manuscripts. What opportunities for training and professional development has the project provided?The grant supported the training of a postdoctoral scholar. The postdoctoral scholar supported by this grant has received a faculty position at UC Riverside How have the results been disseminated to communities of interest? The results were shared through conference presentations and patent application filling. What do you plan to do during the next reporting period to accomplish the goals?Aim 1:Develop and evaluate food-grade photoactive antimicrobial coatings for model zone I and zone II surfaces to reduce the risk of cross-contamination from both bacterial and viral pathogens and bacterial biofilms: In this direction, we aim to develop novel deposition methods that can be adapted to legacy equipment, including evaluating stability of these antimicrobial coatings. Aim 2:Evaluate enhancement in the efficacy of the modified coatings to reduce the risk of cross-contamination by developing affinity-enabled photoactive antimicrobial coatings and photoactive antimicrobial "slippery" coatings with an additional antifouling functionality: Building on the success of the affinity approach, we plan to evaluate the mechanism of this enhancement and use this understanding for broader application of this concept for diverse food and agricultural applications.

Impacts
What was accomplished under these goals? Addressing the challenges of pathogenic microbes requires the development of novel approaches to manage microbial risks in food and agriculture systems. Many alternative solutions based on either bio or chemical approaches lack efficacy compared to conventional antimicrobials in terms of the rate and level of inactivation achieved and thus have not been widely adopted in the industry. Purpose: This research focused on developing novel visible photoactive solid matrices modified with bio-affinity ligands to rapidly inactivate a wide range of target microbes. This approach addresses many of the challenges of conventional photosensitizers and develops novel antimicrobial materials that can be used for diverse applications, including coatings. Method: Curcumin-modified nanofiber membranes and zein membranes were functionalized with yeast cell wall particles as a bio-affinity ligand for capturing diverse pathogens. The increased localization of target microbes in the vicinity of the photoactive species enhanced the antimicrobial activity of generated reactive oxygen species upon exposure to light. The experimental validation of affinity ligands' role in enhancing the antimicrobial performance and rate of inactivation was investigated against inoculated Escherichia coli O157:H7, Listeria innocua, Candida albicans, and bacteriophage T7, respectively. Results: The bio-functionalized matrices showed a high affinity for binding the diverse microbial targets and significant enhancement in generated reactive oxygen species using visible light. This combined effect resulted in a 15 to 30-fold increase in the inactivation rate of tested bacterial, fungal, and viral targets compared to conventional photosensitizer materials. Using visible light-induced ROS, these novel materials achieved more than 7 log (99.99999%) inactivation of the target microbial population in a short treatment time ranging between 2-5 min. These novel materials could be reused for multiple cycles, achieving significant inactivation for at least six consecutive treatment cycles. ?Significance: This work illustrates the development of the next generation of light-induced materials for ultrafast inactivation of diverse microbes. Applications of these materials in food and agricultural systems can enhance food safety and reduce the risk of contamination from food contact surfaces.

Publications

Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Development of visible visible-light-induced antimicrobial materials for ultrafast inactivation of microbes, A. El-Moghazy, N. Nitin, IAFP, 2024

Progress 04/01/22 to 03/31/23

Outputs
Target Audience:The target audience includes researchers at universities, government, and food industries. In addition, the technologies developed through this project are of broader interest to food companies for the translation of research from the lab to industrial practice. Changes/Problems:The project initiation was delayed by six months due to visa delays for a postdoctoral scholar recruited for this project What opportunities for training and professional development has the project provided?The project supported the training of one postdoctoral scholar. How have the results been disseminated to communities of interest?Based on year one results, we have submitted an abstract to the IAFP meeting and are planning to present this to the audience at the annual IAFP meeting What do you plan to do during the next reporting period to accomplish the goals?The plans for the next phase are: 1. Evaluate the anti-viral activity of the Slippery coatings and other food grade component coatings 2.Evaluate enhancement in the efficacy of the modified coatings to reduce the risk of cross-contamination by developing affinity enabled photoactive antimicrobial coatings and photoactive antimicrobial "slippery" coatings with an additional antifouling functionality

Impacts
What was accomplished under these goals? Introduction:Cross-contamination of food with pathogens, including bacteria and viruses, is a major cause of food-borne disease outbreaks across multiple segments of the food industry. The risk of cross-contamination is significant in diverse environments, including harvesting, post-harvest processing, and food service industries. Purpose:This work was focused on the development of antimicrobial "slippery" coatings using photoactive food-grade compositions to reduce biofouling, improve microbial inactivation and provide continuous- and self-sanitation surfaces. Methods:Nanofibrous membranes were functionalized with fluorosilane (approved for food contact surfaces) for reducing surface energy and curcumin for daylight-induced photoactive antimicrobial function. A nanofibers suspension was used as a spray coating on the stainless-steel surface, followed by silicone oil infusion as a lubrication agent to create the antimicrobial slippery surfaces (SLIPS). The antimicrobial activity of the coated surfaces was investigated againstEscherichia coliO157:H7 andListeria innocua. The anti-cross-contamination property of the designed surfaces was examined by conducting a spinach leaf-surface-leaf cross-contamination test. Results:The designed surface showed superhydrophobic properties with a contact angle of 166°. Without the antimicrobial function, SLIPS surface significantly reduced the cross-contamination risk, reducing more than three and four logs of bacterial transfer from the contaminated spinach leaf to the SLIPS surface and the non-infected spinach leaf, respectively. With curcumin modification, the designed surface exhibited excellent self-sanitation property by reduction of 7 log (99.99999%) of both tested microbes after daylight irradiation time of 10 min. After continuous daylight exposure for seven days, the prepared antimicrobial SLIPS exhibited good photostability and retained its light-induced self-sanitation power. Furthermore, the designed surface showed self-sanitation durability by reduction of 7 log ofL. innocuafor five consecutive cycles. Significance:This work illustrates the development of continuous and self-sanitation of surfaces in food processing and food service environments that can reduce the risk of microbial contamination and improve food safety and quality.

Publications

Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: Development of Continuous-and Self-Sanitizing Surface Coatings Based on Visible Light to Prevent Cross-Contamination, A El-Moghazy, N Wisuthiphaet, N Nitin - IAFP 2023, 2023